Antibody Formats

Trends in clinical development of monoclonal antibodies indicates that there is a nascent shift toward the study of antibody fragments and it is likely that antibody fragments are going to be the next important class of protein-based therapeutics after monoclonal antibodies. Antibody fragments [fragments], in particular the scFvs, Fabs and V_HH/V_H retain full antigen-binding capacity and superior properties for research, diagnostic and therapeutic applications. Fragments are particularly useful in applications where epitope binding is sufficient for the desired effect including therapeutic applications such as virus neutralization or receptor blocking.

The smallest antigen binding fragment that retains its complete antigen binding site is the Fv fragment, which consists entirely of variable (V) regions. A soluble and flexible amino acid peptide linker is used to connect the V regions to a scFv (single chain fragment variable) fragment for stabilization of the molecule, or the constant (C) domains are added to the V regions to generate a Fab fragment [fragment, antigen-binding]. scFv and Fab are widely used fragments that can be easily produced in prokaryotic hosts. Other Ab formats include disulfide-bond stabilized scFv (ds-scFv), single chain Fab (scFab), as well as di- and multimeric antibody formats like dia-, tria- and tetra-bodies, or minibodies (miniAbs) that comprise different formats consisting of scFvs linked to oligomerization domains. The smallest fragments are V_HH/V_H of camelid heavy chain Abs and single domain Abs (sdAb). For most therapeutic applications, however, the Fc portion of an Ig is essential as it is instrumental in mediating cytotoxic effector functions such as ADCC and CDC.

Figure 1: The modular domain architecture of Igs has been exploited to create a growing range of alternative antibody formats that spans a molecular-weight range of at least 12–150 kDa and a valency (n) range from monomeric (n = 1), dimeric (n = 2) and trimeric (n = 3) to tetrameric (n = 4) and potentially higher¹. The building block that is most frequently used to create novel antibody formats is the single-chain variable (V)-domain antibody fragment (scFv), which comprises V domains from the heavy and light chain (V_H and V_L domain) joined by a peptide linker of ~15 amino acid residues.

Fab Antibody, scFv, V_HH/V_H Advantages

The previous section in this series is “Antibody Basics”. To review, click here.

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References

1. Carter, P. J. Potent antibody therapeutics by design. Nat Rev Immunol 6, 343-357, doi:10.1038/nri1837 (2006).
2. Yokota, T., Milenic, D. E., Whitlow, M. & Schlom, J. Rapid tumor penetration of a single-chain Fv and comparison with other immunoglobulin forms. Cancer Res 52, 3402-3408 (1992).
3. Jain, R. K. Physiological barriers to delivery of monoclonal antibodies and other macromolecules in tumors. Cancer Res 50, 814s-819s (1990).
4. Ward, E. S., Gussow, D., Griffiths, A. D., Jones, P. T. & Winter, G. Binding activities of a repertoire of single immunoglobulin variable domains secreted from Escherichia coli. Nature 341, 544-546, doi:10.1038/341544a0 (1989).
5. Sanz, L., Cuesta, A. M., Compte, M. & Alvarez-Vallina, L. Antibody engineering: facing new challenges in cancer therapy. Acta Pharmacol Sin 26, 641-648, doi:10.1111/j.1745-7254.2005.00135.x (2005).
6. Holliger, P. & Hudson, P. J. Engineered antibody fragments and the rise of single domains. Nat Biotechnol 23, 1126-1136, doi:10.1038/nbt1142 (2005).
7. Hudson, P. J. & Souriau, C. Engineered antibodies. Nat Med 9, 129-134, doi:10.1038/nm0103-129 (2003).

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